Research in immunology and virology has made enormous progress toward the design of an effective vaccine to prevent Hepatitis C infection, but multidisciplinary efforts are needed to move these designs to the ultimate manufacture of a cost-effective vaccine for global immunization. This proposed Cooperative Research Center (CRC) will critically evaluate a new means to biomanufacture HCV antigens known to induce specific immune responses to HCV - identified through human clinical studies, and predicted to be important in viral clearance - and evaluate responses to these antigens in appropriate in vitro and in vivo model systems. One of the co-Directors of this CRC has lead Chiron's discovery of recombinant antigens from HCV;the evaluation of his currently available suite of proteins in clinical trials indicates that a prophylactic and immunotherapeutic vaccine is feasible. Unfortunately, at least from the standpoint of manufacturing for public health markets, the optimal vaccine should consist of a "cocktail" of multiple antigens to elicit both antibody-mediated and T cell-mediated immune responses. The likely need for two separate protein expression systems to produce this vaccine, one limited by low yields, raises obstacles to commercial development - especially for global use. To overcome these production constraints, this CRC proposes to rapidly utilization current understanding of the properties of important vaccine components to develop and validate a new biomanufacturing system for multiple HCV antigens. A unique feature of this proposed CRC will be the use of green plants as versatile, highly cost-effective production system for recombinant proteins. The research proposed is an interactive partnership that draws upon Chiron's vaccine development expertise to critically evaluate HCV candidate vaccines produced by an academic team that has pioneered the use of plant biotechnology for subunit vaccine manufacture for more than a decade. PROJECT 1: Plant- Derived Envelope Glycoprotein E1/E2 Vaccine for HCV (Mason, H.) PROJECT 1 DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) is a major worldwide agent of acute and chronic hepatitis, with over 3% of the world's population infected and at risk for liver cirrhosis and cancer, and a US cost burden of $1 billion per year. Studies showed that vaccine development should target both antibody and T-cell mediated immunity. Recombinant envelope glycoproteins gpE1 and gpE2 are the leading vaccine candidates for antibody-mediated protection, showing protection in chimpanzee. However, gpE1/gpE2 is currently produced in cultured mammalian cells, with a limited capacity for scale-up to achieve global immunization. Alternatively, production of gpE1/gpE2 in plant systems has the potential for vast scale-up. We hypothesize that: 1) HCV gpE1/gpE2 vaccine can be produced in a plant system in a form that is equivalent to the mammalian cell product used for injection;2) HCV gpE1/gpE2 vaccine can be scaled up to a commercially competitive level using a leaf or a grain seed-based expression system;and 3) Plant-derived HCV gpE1/gpE2 can stimulate antibody responses when orally delivered by ingestion of minimally processed plant tissue. The specific aims of this proposal will address these questions by using plant-optimized gpE1/gpE2 genes expressed in plants to assess the potential to assemble immunogenic gpE1/gpE2 heterodimers. We will examine tobacco leaf and corn seed expression systems and evaluate gpE1/gpE2 accumulation and quality to select the optimal system. M. Houghton at Chiron Corporation leads their HCV vaccine effort and will direct Project 3 of this HCV Research Center, which will purify gpE1/gpE2 from plant sources and test its immunogenic capacity in animal models. We will furthermore test the potential for use of orally delivered plant-derived gpE1/gpE2 vaccine, with the aim to develop a convenient and economical vaccine for the developing world. If successful, these studies will greatly enhance the capacity to produce an injectable vaccine for HCV, while also offering the possibility to improve worldwide immunization coverage with an oral vaccine.